Disclosure of Invention
An embodiment of the present invention provides a surface plasmon resonance system, and aims to solve the problem that the dynamic range of the existing surface plasmon resonance system is small.
The embodiment of the present invention is achieved as follows, and a surface plasmon resonance system includes:
a line light source;
a polarizer for obtaining polarized light from the linear light source;
the optical modulator is used for changing the spatial phase distribution of the polarized light to form modulated polarized light;
a beam splitter for splitting the modulated polarized light into probe light and reference light;
a converging element for converging the probe light into a point;
the prism is provided with a sample cell and is used for receiving the detection light and enabling a convergence point of the detection light to be positioned on the sensing film;
the polarization direction of the first analyzer is vertical to that of the polarizer, and the first analyzer is matched with the optical modulator to periodically modulate the probe light;
a dispersion element for decomposing the probe light to form a spectrum;
the area array detector is used for recording the intensity of the spectrum so as to form a detection image which is simultaneously resolved by an incident angle and a spectrum wavelength;
the polarization direction of the second analyzer is vertical to that of the polarizer, and the second analyzer is matched with the optical modulator to perform periodic modulation on the reference light;
a photodetector for converting the reference light into a reference electrical signal; and
and the computer is used for acquiring and analyzing the detection image and the reference electric signal and controlling the optical modulator.
It is another object of an embodiment of the present invention to provide a method for detecting by using the above surface plasmon resonance system, the method comprising the steps of:
illuminating the line light source, injecting a first detection sample into the sample cell, and acquiring a series of first detection images with incident angles and spectral wavelengths resolved simultaneously by the area array detector at equal time intervals or unequal time intervals in one modulation period of the modulated polarized light;
injecting a second detection sample into the sample cell, enabling the second detection sample to react with the first detection sample, and acquiring a series of second detection images with incident angles and spectral wavelengths resolved simultaneously by the area array detector at equal time intervals or unequal time intervals in one modulation period of the modulated polarized light;
and searching pixels which generate the best SPR phenomenon in the first detection image and the second detection image so as to calculate the refractive index change quantity delta n' of the first detection sample or the second detection sample, thereby obtaining the interaction between the first detection sample and the second detection sample.
The embodiment of the invention firstly obtains polarized light from a linear light source and converts the polarized light into modulated polarized light, then the modulated polarized light is divided into detection light and reference light, the detection light is converged on a sensing film of a prism, the detection light at the moment simultaneously comprises incident angle information and spectral wavelength information, the intensity of a spectrum formed after the detection light emitted from the prism is dispersed is recorded by an area array detector, and a detection image with the incident angle and the spectral wavelength being simultaneously resolved is formed. The detection image formed by the method is a three-dimensional image, wherein one dimension comprises angle information, namely, each pixel of the area array detector in the direction parallel to the linear light source represents different light incidence angles; the other dimension contains wavelength information, namely, each pixel of the area array detector in the direction vertical to the linear light source (also in the spectral direction) represents different light wavelength; the other dimension contains phase information, i.e. the phase change of the probe light relative to the reference light (also SPR phase). In a modulation period of the modulated polarized light, the area array detector acquires a series of detection images at equal time intervals or unequal time intervals, the light intensity recorded by the pixels at the same position in the series of images changes with time to form a curve, the area array detector is provided with a plurality of pixels, a plurality of curves can be formed, each curve contains SPR phase information, the phase of each curve is obtained through calculation, then the phase difference between each curve and the reference electric signal is calculated finally by combining the reference electric signal generated by the photoelectric detector, and the change of the phase difference reflects the change of the refractive index of the detection sample.
During testing, the line light source is firstly lightened, a first detection sample is injected into the sample cell, and a series of first detection images which are simultaneously resolved by the incident angle and the spectral wavelength are obtained by the area array detector in a mode of equal time interval or unequal time interval in one modulation period of the modulated polarized light. And then injecting a second detection sample into the sample cell to react with the first detection sample, and acquiring a series of second detection images resolved by the incident angle and the spectral wavelength at the same time by the area array detector at equal time intervals or at unequal time intervals in one modulation period of the modulated polarized light. And then searching pixels which are optimal for generating an SPR phenomenon in the first detection image and the second detection image so as to calculate the refractive index change quantity delta n' of the first detection sample or the second detection sample, thereby acquiring the interaction between the first detection sample and the second detection sample. The measurement of the refractive index change in a large range is carried out by searching for the pixel which best generates the SPR phenomenon in the first detection image and the second detection image, namely, the phase SPR high-sensitivity detection, and the first detection image and the second detection image are three-dimensional images of an SPR phase, an incidence angle and a spectral wavelength, and the dynamic range of the three-dimensional images is large, so that the application requirement of the surface plasma resonance system with a large dynamic range is met, and the high sensitivity is also guaranteed.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
The embodiment of the invention firstly obtains polarized light from a linear light source and converts the polarized light into modulated polarized light, then the modulated polarized light is divided into detection light and reference light, the detection light is converged on a sensing film of a prism, the detection light at the moment simultaneously comprises incident angle information and spectral wavelength information, the intensity of a spectrum formed after the detection light emitted from the prism is dispersed is recorded by an area array detector, and a detection image with the incident angle and the spectral wavelength being simultaneously resolved is formed. The detection image formed by the method is a three-dimensional image, wherein one dimension comprises angle information, namely, each pixel of the area array detector in the direction parallel to the linear light source represents different light incidence angles; the other dimension contains wavelength information, namely, each pixel of the area array detector in the direction vertical to the linear light source (also in the spectral direction) represents different light wavelength; the other dimension contains phase information, i.e. the phase change of the probe light relative to the reference light (also SPR phase). In a modulation period of the modulated polarized light, the area array detector acquires a series of detection images at equal time intervals or unequal time intervals, the light intensity recorded by the pixels at the same position in the series of images changes with time to form a curve, the area array detector is provided with a plurality of pixels, a plurality of curves can be formed, each curve contains SPR phase information, the phase of each curve is obtained through calculation, then the phase difference between each curve and the reference electric signal is calculated finally by combining the reference electric signal generated by the photoelectric detector, and the change of the phase difference reflects the change of the refractive index of the detection sample.
During testing, the line light source is firstly lightened, a first detection sample is injected into the sample cell, and a series of first detection images which are simultaneously resolved by the incident angle and the spectral wavelength are obtained by the area array detector in a mode of equal time interval or unequal time interval in one modulation period of the modulated polarized light. And then injecting a second detection sample into the sample cell to react with the first detection sample, and acquiring a series of second detection images resolved by the incident angle and the spectral wavelength at the same time by the area array detector at equal time intervals or at unequal time intervals in one modulation period of the modulated polarized light. And then searching pixels which are optimal for generating an SPR phenomenon in the first detection image and the second detection image so as to calculate the refractive index change quantity delta n' of the first detection sample or the second detection sample, thereby acquiring the interaction between the first detection sample and the second detection sample. The measurement of the refractive index change in a large range is carried out by searching for the pixel which best generates the SPR phenomenon in the first detection image and the second detection image, namely, the phase SPR high-sensitivity detection, and the first detection image and the second detection image are three-dimensional images of an SPR phase, an incidence angle and a spectral wavelength, and the dynamic range of the three-dimensional images is large, so that the application requirement of the surface plasma resonance system with a large dynamic range is met, and the high sensitivity is also guaranteed.
The following describes the implementation of the present invention in detail with reference to specific embodiments.
As shown in fig. 1, the surface plasmon resonance system provided by the embodiment of the invention is composed of a probe optical path and a reference optical path. The detection light path comprises a linear light source 1, a polarizer 2, an optical modulator 3, a beam splitter 4, a converging element 5, a prism 6 provided with a sample cell 60, a first analyzer 7, a dispersion element 8 and an area array detector 9. The reference light path comprises a linear light source 1, a polarizer 2, a light modulator 3, a beam splitter 4, a second analyzer 10 and a photoelectric detector 11.
The linear light source 1, the polarizer 2, the optical modulator 3 and the beam splitter 4 are shared by a detection light path and a reference light path. The line light source 1 is here a quasi-linear light source, which is formed by a broadband light source 12 (such as an LED, a white light source, etc.) via an aspherical lens 13 (such as a collimating lens) and a slit 14. Here, the polarizer 2 obtains polarized light required for the present embodiment from the linear light source 1. The spatial phase distribution of the polarized light is changed by the optical modulator 3 to become modulated polarized light. The modulated polarized light is split into probe light and reference light by a beam splitter 4. The probe and reference optical paths are described in detail below.
In the embodiment of the present invention, the converging element 5 receives the probe light, and the converging element 5 is a first cylindrical mirror that converges the probe light into a point. The probe light is received by the prism 6 provided with the sample cell 60 and has its point of convergence at the sensing film 61, where the probe light contains both incident angle information and spectral wavelength information. The detection light reflected by the sensing film 61 is projected to a first analyzer 7, the polarization direction of the first analyzer 7 is perpendicular to the polarization direction of the polarizer 2, and the detection light is periodically modulated by matching with the light modulator 3. The detection light emitted through the prism 6 and the first analyzer 7 is linear detection light. At this time, the probe light is projected to a dispersion element 8 (such as a prism, a grating, etc.), and the probe light is decomposed by the dispersion element 8 to form a spectrum. Finally, the intensity of the spectrum is recorded by an area array detector 9 (such as a CCD, a CMOS and the like), so that a detection image with the incident angle and the spectrum wavelength resolved simultaneously is formed.
The detection image thus formed is a three-dimensional image, wherein one dimension contains angle information, i.e. each pixel of the area array detector 9 in the direction parallel to the line light source represents a different light incident angle; the other dimension contains wavelength information, that is, each pixel of the area array detector 9 in the direction perpendicular to the line light source (also referred to as the spectral direction) represents different light wavelength; the other dimension contains phase information, i.e. the phase change of the probe light relative to the reference light (also SPR phase). In a modulation period of the modulated polarized light, the area array detector 9 acquires a series of detection images at equal time intervals or unequal time intervals, the light intensity recorded by the same pixel in the series of images changes with time to form a curve, the area array detector 9 is provided with a plurality of pixels to form a plurality of curves, each curve contains SPR phase information, the phase of each curve can be obtained through calculation, then the phase difference between each curve and a reference electric signal is calculated by combining a reference electric signal, namely a sine or cosine reference signal, generated by the photoelectric detector 11, and the change of the phase difference reflects the change of the refractive index of a detection sample.
In the embodiment of the present invention, the reference light is received by the second analyzer 10, and the polarization direction of the second analyzer 10 is perpendicular to the polarization direction of the polarizer 2, and is matched with the optical modulator 3 to perform periodic modulation on the reference light. The reference light is then converted into a reference electrical signal by a photodetector 11 (e.g., PIN, photocell, etc.), which is typically collected by a data acquisition card 15.
In the embodiment of the present invention, the computer 16 collects and analyzes the detection image and the reference electrical signal, and controls the optical modulator 3 to periodically modulate the polarized light. The computer 16 generally outputs a sine or cosine spatial light modulation signal to the light modulator 3, so that the light intensity curve recorded by the same pixel in the multiple detected images is a sine or cosine curve, as shown in fig. 2 and 3. Here the computer 16 may also cause sine or cosine spatial light modulation signals to be output to the light modulator 3 via a data acquisition card 15.
The detection beam emitted from the prism 6 is wide, and the detection area of the existing area array detector 9 is small. A beam shrinking device 17 for shrinking the detection light to a size corresponding to the area array detector 9 is arranged between the prism 6 and the first analyzer 7. The beam reducer 17 is composed of a second cylindrical mirror 18, a third cylindrical mirror 19, and a collimator lens 20, which are arranged in this order along the optical axis.
In the embodiment of the invention, the modulated polarized light is partially reflected by the beam splitter 4 to form the reference light, and the modulated polarized light is partially transmitted by the beam splitter 4 to form the detection light. The dispersive element is a dispersive prism 8, and the detection light emitted by the first analyzer 7 is parallel to the bottom surface 81 of the dispersive prism.
In summary, since each pixel corresponds to a different incident angle in the direction of the parallel light source, the phase change at each incident angle can be obtained; meanwhile, since each pixel corresponds to a different wavelength in a direction perpendicular to the line light source, a phase change at each wavelength can be obtained. By combining the incident angle and wavelength information, SPR phase information at the optimal incident angle and wavelength can be obtained, wherein the SPR information is most sensitive. That is, a three-dimensional composite image is formed by three variables of angle, wavelength and phase, where the X-axis is the angle (or wavelength), the Y-axis is the wavelength (or angle), and the Z-axis is the phase change of the probe light with respect to the reference light (i.e., SPR phase). When the refractive index of the sample changes, there is a position (here, represented by a pixel) where the phase change is the largest in the three-dimensional map, and the refractive index change of the detection sample can be calculated from the phase change of the detection light corresponding to the pixel. Therefore, the embodiment of the invention has a large dynamic range, and more importantly, the wavelength and the incident angle are not fixed values adopted by the common SPR technology, but wide incident angle and spectral wavelength resolution detection are simultaneously carried out in parallel in a large range, and no matter how large the refractive index changes, the optimal incident angle and spectral wavelength can be always ensured to generate the optimal SPR phenomenon. Theoretically, the dynamic range can be infinitely large, and is limited by the angle of the convergent light of the cylindrical mirror and the spectral width of the light source in practice.
When the system is used for detection, the line light source is firstly lightened, a first detection sample is injected into the sample cell, and a series of first detection images which are simultaneously resolved by the incident angle and the spectral wavelength are obtained by the area array detector in a mode of equal time interval or unequal time interval in one modulation period of the modulated polarized light. And then injecting a second detection sample into the sample cell to react with the first detection sample, and acquiring a series of second detection images resolved by the incident angle and the spectral wavelength at the same time by the area array detector at equal time intervals or at unequal time intervals in one modulation period of the modulated polarized light. And then searching pixels which are optimal for generating an SPR phenomenon in the first detection image and the second detection image so as to calculate the refractive index change quantity delta n' of the first detection sample or the second detection sample, thereby acquiring the interaction between the first detection sample and the second detection sample.
The step of finding the best pixel generating the SPR phenomenon in the first and second detected images is specifically: extracting the phase of the detection light corresponding to each pixel of the first detection image, comparing the phase with the phase of the reference light, calculating the difference between the phase of the detection light corresponding to each pixel of the first detection image and the phase of the reference light, and recording the phase difference as the initial phase of the first detection sample before reaction with the second detection sample; extracting the phase of the detection light corresponding to each pixel of the second detection image, comparing the phase with the phase of the reference light, calculating the difference between the phase of the detection light corresponding to each pixel of the second detection image and the phase of the reference light, and recording the phase difference as the change phase after the reaction of the first detection sample and the second detection sample; and calculating the difference between the initial phase and the change phase of the corresponding pixel in the first detection image and the second detection image, recording the difference as the action phase before and after the reaction of the first detection sample and the second detection sample, and selecting the pixel corresponding to the maximum absolute value of the action phase, namely the pixel with the best SPR phenomenon generated in the first detection image and the second detection image.
The pixels of the area array detector 9 can be represented by λ and θ, where λ represents the wavelength of the light incident on the pixel, and θ represents the incident angle of the light incident on the pixel when the light is incident on the sensing film. For example, λ can be used for the first pixel on the area array detector 91、θ1Indicating that the second pixel can be represented by2、θ2Indicating that the third pixel can be represented by3、θ3Is said. It should be understood that the pixels of the area array detector 9 correspond one-to-one to the pixels of the detected image.
According to the phase shift formula of the phase type SPR, the phase curve of the SPR at different angles and wavelengths is made in the embodiment of the invention, as shown in FIG. 4. The phase shift formula of the phase type SPR here is:
<math>
<mrow>
<mover>
<msub>
<mi>r</mi>
<mi>p</mi>
</msub>
<mo>~</mo>
</mover>
<mo>=</mo>
<mfrac>
<mrow>
<msub>
<mi>r</mi>
<mn>01</mn>
</msub>
<mo>+</mo>
<msub>
<mi>r</mi>
<mn>12</mn>
</msub>
<msup>
<mi>e</mi>
<mrow>
<mn>2</mn>
<mi>i</mi>
<msub>
<mi>k</mi>
<mrow>
<mn>1</mn>
<mi>z</mi>
</mrow>
</msub>
<mi>d</mi>
</mrow>
</msup>
</mrow>
<mrow>
<mn>1</mn>
<mo>+</mo>
<msub>
<mi>r</mi>
<mn>01</mn>
</msub>
<mo>·</mo>
<msub>
<mi>r</mi>
<mn>12</mn>
</msub>
<msup>
<mi>e</mi>
<mrow>
<mn>2</mn>
<mi>i</mi>
<msub>
<mi>k</mi>
<mrow>
<mn>1</mn>
<mi>z</mi>
</mrow>
</msub>
<mi>d</mi>
</mrow>
</msup>
</mrow>
</mfrac>
<mo>-</mo>
<mo>-</mo>
<mo>-</mo>
<mrow>
<mo>(</mo>
<mn>1</mn>
<mo>)</mo>
</mrow>
</mrow>
</math>
<math>
<mrow>
<msub>
<mi>X</mi>
<mi>j</mi>
</msub>
<mo>=</mo>
<mfrac>
<msub>
<mi>ϵ</mi>
<mi>j</mi>
</msub>
<msub>
<mi>k</mi>
<mi>jz</mi>
</msub>
</mfrac>
<mo>-</mo>
<mo>-</mo>
<mo>-</mo>
<mrow>
<mo>(</mo>
<mn>3</mn>
<mo>)</mo>
</mrow>
</mrow>
</math>
<math>
<mrow>
<msub>
<mi>k</mi>
<mi>jz</mi>
</msub>
<mo>=</mo>
<mfrac>
<mi>ω</mi>
<mi>c</mi>
</mfrac>
<msqrt>
<msub>
<mi>ϵ</mi>
<mi>j</mi>
</msub>
<mo>-</mo>
<msub>
<mi>ϵ</mi>
<mn>0</mn>
</msub>
<msup>
<mi>sin</mi>
<mn>2</mn>
</msup>
<mi>θ</mi>
</msqrt>
<mo>,</mo>
<mi>j</mi>
<mo>=</mo>
<mn>0,1,2</mn>
<mo>-</mo>
<mo>-</mo>
<mo>-</mo>
<mrow>
<mo>(</mo>
<mn>4</mn>
<mo>)</mo>
</mrow>
</mrow>
</math>
wherein,is a reflection coefficient, ri,i+1(i is 0, 1) is the interface reflection coefficient, d is the thickness of the metal film, ω is the angular frequency of the incident light, c is the speed of light in vacuum,j(j is 0, 1, 2) is the dielectric constant of the prism, the metal film and the dielectric, respectively, kjz(j is 0, 1, 2) wave vectors of the prism, the metal film and the dielectric respectively, theta is an incident angle of the light wave in the incident medium,is the phase of the optical wave.
According to the embodiment of the invention, the refractive index change delta n' of the first detection sample or the second detection sample can be calculated according to the SPR phase curves under different incidence angles and wavelengths. For example, finding out in the first and second detected imagesThe pixel best for generating the SPR phenomenon is the first pixel (lambda) of the area array detector 91、θ1) At this time, the first pixel (λ) is selected1、θ1) And calculating the refractive index change delta n' of the first detection sample or the second detection sample according to the corresponding SPR phase curve. As another example, the pixel best for generating the SPR phenomenon is found in the first detection image and the second detection image as the third pixel (λ) of the area array detector 93、θ3) At this time, the third pixel (λ)3、θ3) And calculating the refractive index change delta n' of the first detection sample or the second detection sample according to the corresponding SPR phase curve. Of course, if the first probe sample is DNA, then the second probe sample is protein; if the first test sample is an antigen, the second test sample is an antibody.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are intended to be included within the scope of the present invention.